A large number of mitochondrial mutants of the human cell line VA2-B have been recently isolated by using cells partially depleted of their mitochondrial DNA (mtDNA) and by using specific mutagenic treatments designed to enhance the recovery of mtDNA mutations. These mutants include clones resistant to inhibitors of mitochondrial respiration and oxidative phosphorylation, and clones deficient in mitochondrial respiratory function. Several of these mutants are specifically deficient in mitochondrial protein synthesis (mtPS minus) and the genes coding for these defects have been localized on mtDNA. We are presently analyzing these mutants in detail biochemically in order to determine if mitochondrial transcription or ribosomal biogenesis are affected. Furthermore we are studying the mtDNA sequence organization of selected mutants by gel electrophoresis of restriction digests to detect potential sequence alterations, such as deletions and also are determining the mtDNA content of the mutant cells. In addition, the residual products of mitochondrial protein synthesis in mitochondrial protein synthesis deficient mutants, if any, are being analyzed by gel electrophoresis and the ultrastructural characteristics of their mitochondria are being analyzed in the electron microscope. Additional genetic studies of these mtPS minus mutants are in progress and include complementation analysis in cell hybrids and the isolation of revertants and potential suppressors. Another area of study includes the ability of potential mtDNA specific mutagens to induce new mtDNA mutants.